For men in the U.S., prostate cancer is the most commonly diagnosed cancer, and the second leading cause of cancer-related death (Greenlee R T, Murray T. Bolden S. Wingo P A. Cancer statistics, 1999. Ca: a Cancer Journal for Clinicians 2000:50:7-33). Prostate cancers originate as localized lesions; some of these localized lesions will progress to become invasive, migratory and metastatic. Our current understanding of the mechanisms of the prostate cancer invasion process, however, is poor. Our ability to predict the acquisition of invasive potentials by a prostate cancer is limited.
The mechanisms leading to the development of a prostate cancer are complex. Currently, it is believed to be the result of multiple transformation steps from normal prostate glandular cells (Carter H B. Piantadosi S. Isaacs J T. Clinical evidence for and implications of the multistep development of prostate cancer. Journal of Urology. 143(4):742-6, 1990). The initial steps result in what are described as prostatic interepithelial neoplastic (PIN) lesions (Isaacs J T. Molecular markers for prostate cancer metastasis. Developing diagnostic methods for predicting the aggressiveness of prostate cancer. [Review] [92 refs] American Journal of Pathology. 150(5):1511-21, 1997). These PIN lesions may then typically have three different fates based on an assessment of their impact to the patient. The PIN lesions can remain as such, not producing histologically detectable prostate cancer, or further transform into histologically detectable prostate cancer. Most of the histologically detectable prostate cancers will be asymptomatic in the patient and remain non-manifest clinically as many are discovered post-mortem (Carter H Coffey D. Prostate Cancer: the magnitude of the problem in the United States. In a Multidisciplinary Analysis of Controversies in the Management of Prostate Cancer. (Eds. Coffey D. Resnick M. Door R. et al.), pp1-9, Plenum Press, 1988; Carter H B Piantadosi S. Issacs J T. Clinical evidence for implications of the multistep development of prostate cancer. Journal of Urology. 143(4):742-6, 1990; Scardino P T. Weaver R. Hudson M A. Early detection of prostate cancer. [Review] [102 refs] Human Pathology. 23(3):211-22, 1999). Prostate cancers are diagnosed clinically by an estimate of size and location using the TNM staging system (Denis L J. Staging and prognosis of prostate cancer. European Urology. 24 Suppl 2:13-8, 1993), and by pathological staging based on an examination of the histology of the removed prostate via either biopsy or prostatectomy using a system by D. F. Gleason, (Gleason D F. Classification of prostatic carcinomas. Cancer Chemotherapy Reports-Part 1. 50(3):125-8, 1966). About 50% of prostate cancer cases receiving treatment are diagnosed clinically as advanced, or, non-organ-confined (Scardino P T. Weaver R. Hudson M A. Early detection of prostate cancer. [Review] [102 refs] Human Pathology. 23(3):211-22, 1992), for which no effective treatment exists (Yagoda A. Petrylak D. Cytotoxic chemotherapy for advanced hormone-resistant prostate cancer. [Review] [63 refs] Cancer. 71(3 Suppl): 1098-109, 1993 and Petrylak, 1993). Of the remaining 50% cases, ⅓ (xcx9c50,000) are diagnosed as organ-confined but micrometastasis may be present. The final group of patients (xcx9c100,000) have truly organ-confined prostate cancer and can be cured by radical prostatectomy (Sgrignoli A R Walsh P C. Steinberg G D. Steiner M S. Epstein J I. Prognostic factors in men with stage D1 prostate cancer: identification of patients less likely to have prolonged survival after radical prostatectomy [see comments]. Journal of Urology. 152(4):1077-81, 1994; Zincke H. Oesterling J E. Blute M L. Bergstralh E J. Myers R P. Barrett D M. long term (15 years) results after radical prostatectomy for clinically localized (stage T2c or lower) prostate cancer [see comments]. Journal of Urology. 152(5 Pt 2): 1850-7, 1994) or left untreated (watchful waiting) without the risk of life-threatening or life-altering. For the patients with non-organ-confined prostate cancers (discovered via either biopsy or surgery), undergoing systemic treatment early is essential to the management of their cancer (Yagoda A. Petrylak D. Cytotixic chemotherapy for advanced hormone-resistant prostate cancer [Review] [63 refs] Cancer. 71(3 Suppl): 1098-109, 1993). Consequently, it will be ideal, both medically and economically, if one could precisely predict upon early pathological examination of the tumor, which group of patients will have truly organ-confined disease versus which group will have invasive prostate cancer.
Clinical staging of prostate cancer generally depends on the results of three tests that are performed in the following order: a PSA (prostate-specific antigen) blood test as a screening method; DRE (digital rectal examination) for an initial indication of palpable disease; and, a biopsy to obtain samples for histological examination. Prostate cancers, removed either via biopsy or surgery, are graded histologically by the system of Gleason. (Gleason D F. Classification of prostatic carcinomas. Cancer Chemotherapy Reportsxe2x80x94Part 1. 50(3):125-8, 1966), which is an evaluation of how aggressive and how poorly-differentiated the prostate cancers are. The aggressiveness of prostate tumors: of low Gleason scores ( less than 5) is limited; of high Gleason scores (8-10) are highly aggressive; but, for the intermediate Gleason-score (5-7) prostate cancers (76% of prostate tumors), the accuracy of predicting their aggressiveness is poor (Gleason D F. Mellinger G T. Prediction of prognosis for prostatic adenocarcinoma by combined histological grading and clinical staging. Journal of Urology. 111(1):58-64, 1974). Thus, the ability to accurately determine the aggressiveness of these intermediate Gleason-score prostate tumors has remained as a practical challenge to, and a primary goal for, prostate cancer research (Isaacs J T. Molecular markers for prostate cancer metastasis. Developing diagnostic methods for predicting the aggressiveness of prostate cancer. [Review] [92 refs] American Journal of Pathology. 150(5):1511-21, 1997). Especially with regard to the number of patients (150,000) facing a decision of whether to undergo systemic treatment, the most urgent demand in prostate cancer care is the development of methods to enhance our ability to accurately predict the aggressiveness of the tumors with Gleason scores of 5-7.
It is now commonly believed that cancers occur via multiple transformation steps by accumulating mutations in three classes of genes: proto-oncogenes (Park M. Oncogenes. In The Genetic Basis of Human Cancer (Eds. Vogelstein B and Kinzler K W). pp205-28. McGraw-Hill Health Professions Divisions, 1998); tumor-suppressor genes (Knuutila S. Aalto Y. Bjorkqvist A M. EL-Rifai W. Hemmer S. Huhta T. Kettunen E. Kiuru-Kuhlefelt S. Larramendy M L. Lushnikova T. Monni O. Pere H. Tapper J. Tarkkanen M. Varis A. Wasenius V M. Wolf M. Zhu Y. DNA copy number losses in human neoplasms. [Review] [197 refs] American Journal of Pathology. 155(3):683-94, 1999); and, DNA repair genes (Knuutila S. Aslto Y. Bjorkqvit A M. EL-Rifai W. Hemmer S. Huhta T. Kettunen E. Kiuru-Kuhlefelt S. Larramedy M L. Lushnikova T. Monni O. Pere H. Tapper J. Tarkkanen M. Varis A. Wasenius V M. Wolf M. Zhu Y. DNA copy number losses in human neoplasms. [Review] [197 refs] American Journal of Pathology. 155(3):683-94, 1999). The histological prostate cancers for which the prediction of clinical aggressiveness is difficult (those with the intermediate Gleason scores 5-7) probably have not gone through the necessary xe2x80x9cmulti-stepxe2x80x9d transformation to acquire the potentials to behave aggressively (as would the high-grade cancers). This notion was supported by studies comparing the course of prostate cancer development among men in Japan and in the U.S., and Japanese men who migrated to the U.S. (Carter H B. Piantadosi S. Isaacs J T. Clinical evidence for and implications of the multistep development of prostate cancer. Journal of Urology, 1990; Haenszel W. Kurihara M. Studies of Japanese Migrants. I. Mortality form cancer and other diseases among Japanese in the United States. Journal of the National Cancer Institute. 40(1):43-68, 1968: Akazaki K. Stemmerman G N. Comparative study of latent carcinoma of the prostate among Japanese in Japan and Hawaii. Journal of the National Cancer Institute. 50(5):1137-44, 1973; Dunn J E. Cancer epidemiology in populations of the United Statesxe2x80x94with emphasis on Hawaii and Californiaxe2x80x94and Japan. Cancer Research. 35(11 Pt.2):3240-5, 1975). The findings were that first- and second-generation Japanese men who migrated to the U.S. have a higher prostate cancer incident rate than native Japanese men. The emigrant Japanese men""s prostate cancer incident rate is similar to that of men in the U.S. Investigating the changes of expression in these three classes of cancer-relevant genes during the course of prostate cancer development will lead to a better understanding of the processes by which prostate cancels acquire their aggressive potentials. The discovery of molecules whose changes can be correlated to the staging of prostate cancer will provide new tools to improve our ability to better predict the aggressive behaviors of prostate cancer. This approach is now commonly referred to as xe2x80x9cmolecular stagingxe2x80x9d.
Down-regulated genes such as tumor suppressors, invasion suppressors or metastasis suppressors may be used as prostate cancer markers. Examples of these genes that can potentially serve as prostate cancer markers for xe2x80x9cmolecular stagingxe2x80x9d are: KAI1 (Dong J T. Suzuki H. Pin S S. Bova G S., Schalken J A, Issacs W B, Barrett J C. Issace I T. Down-regulation of the KAI1 metastasis suppressor gene during the progression of human prostatic cancer infrequently involves gene mutation or allelic loss. Cancer Research. 56(19): 4387-90, 1996); (Ueda T, Ichikawa T., Tamaru j, Mikata A, Akakura K, Akimoto S, Imai T. Yoshie O. Shiraishi T. Yatani R. Ito H. Shimazaki J., Expression fof the KAI1 protein in benign prostatic hyperplasia and prostate cancer. American Journal of Pathology 149(5): 1435-40, 1996); E-cadherin (Umbas R. Isaacs W B Bringuier P P. Schaafsma H E. Karthaus H F. Oosterhof G O. Debruyne F M. Schalken J A. Decreased E-Cadherin expression is associated with poor prognosis is patients with prostate cancer. Cancer Research. 52(18):5104-9, 1992, Umbas R. Isaacs W B. Bringuier P P. Schaafsma H E. Karthaus H F. Oosterhof G O Debruyne F M. Schalken J A. Decreased E-cadherin expression is associated with poor prognosis in patients with prostate cancer. Cancer Research. 54(14):3929-33, 1994); xcex21C integrin (Fornaro M. Tallini G. Bofetiado C J. Bosari S. Languino L R. Down-regulation of xcex21C integrin, an inhibitor of cell proliferation, in prostate carcinoma. American Journal of Pathology. 149(3):765-73, 1996 Fornaro M. Manzotti M. Tallini G. Slear A E. Bosari S. Ruoslahti E. Languino L R. xcex21C integrin in epithelial cells correlates with a nonproliferative phenotype: forced expression of xcex21C inhibits prostate epithelial cell proliferation. American Journal of Pathology. 153(4):1079-87, 1998; p27(kip1) (Tsihlias J. Kapusta L R. DeBoer G. Morava-Protzner I. Zbieranowski I. Bhattachaya N. Catzavelos G C. Klotz L H. Slingerland J M. Loss of cyclin-dependent kinase inhibitor p27Kipi is a novel prognostic factor in localized human prostate adenocarcinoma. Cancer Research. 58(3):542-8, 1998); and CD44 (Lou W. Krill D. Dhir R. Becich M J. Dong J T. Frierson H F Jr. Isaacs W B. Isaacs J T. Gao A C. Methylation of the CD44 metastasis suppressor gene in human prostrate cancer. Cancer Research. 59(10):2329-31, 1999). By using the method of immunohistochemistry, these genes were found to be down-regulated in prostate cancer. KAI""s down regulation is a potential predictor of metastasis (Dong, et al., 1996: Ueda et al., 1996). While E-cadherin""s down regulation is strongly correlated to higher Gleason grades. Functional studies of these genes have given clues to their role in prostate cancer or normal prostate biology. (Debruyne F M. Isaacs W B. Expression of the cellular adhesion molecule E-cadherin is reduced or absent in high-grade prostate cancer. Cancer Research. 52(18):5104-9, 1992; Umbas R. Isaacs W B. Bringuier P P. Schaafsma H E. Karthaus H F. Oosterhof G O Debruyne F M. Schalken J A. Decreased E-cadherin expression is associated with poor prognosis in patients with prostate cancer. Cancer Research. 54(14):3929-33, 1994; metastasis suppressor gene during the progression of human prostatic cancer infrequently involves gene mutation or allelic loss. Cancer Research. 56(19):4387-90, 1996 Ueda T. Ichikawa T. Tamaru J. Mikata A. Akakura K. Akimoto S. Imai T. Yoshie O. Shiraishi T. Yatani R. Ito H. Shimazaki J. Expression of the KAI1 protein in benign prostatic hyperplasia and prostate cancer. American Journal of Pathology. 149(5):1435-40, 1996) while E-cadherin""s down-regulation is strongly correlated to higher Gleason grades (Umbas R. Schalken J A. Aalders T W. Carter B S. Karthaus H F. Schaafsma H E. Debruyne F M. Isaacs W B. Expression of the cellular adhesion molecule E-cadherin is reduced or absent in high-grade prostate cancer. Cancer Research. 52(18):5104-9, 1992; Umbas R. Isaacs W B. Bringuier P P. Schaafsma H E. Karthaus H F. Oosterhof G O. Debruyne F M. Schalken J A. Decreased E-cadherin expression is associated with poor prognosis in patients with prostate cancer. Cancer Research. 54(14):3929-33, 1994) Functional studies of these genes have given clues to their role in prostate cancer or normal prostate biology. Human KAI suppress metastasis of rat prostate cancer cells upon gene transfer (Dong J T. Lamb P W. Rinker-Schaeffer C W. Vukanovic J. Ichikawa T. Isaacs J T. Barrett J C. KAI1, a metastasis suppressor gene for prostate cancer on human chromosome 11p11.2 [see comments]. Science. 268(5212):884-6, 1995). xcex21C and p27(kip1) are involved in signaling pathway that inhibits cell proliferation (Fornaro M. Tallini G. Zheng D Q. Flanagan W M. Manzotti M. Languino L R. p27(kip1) acts as a downstream effector of and is coexpressed with the beta1C integrin in prostatic adenocarcinoma. Journal of Clinical Investigation. 103(3):321-9, 1999). E-cadherin expression is progressively lost during the transformation of rat prostate cancer from non-invasive to invasive (Bussemakers M J. van Moorselaar R J. Giroldi L A. Ichikawa T. Isaacs J T. Takeichi M. Debruyne F M. Schalken J A. Decreased expression of E-cadherin in the progression of rat prostatic cancer. Cancer Research. 52(10):2916-22, 1992), and it has also been shown to be an invasion suppressor (Lou W. Krill D. Dhir R. Becich M J. Dong J T. Frierson H F Jr. Isaacs W B. Isaacs J T. Gao A C. Methylation of the CD44 metastasis suppressor gene in human prostate cancer. Cancer Research. 59(10):2329-31, 1999). CD44 loss of expression is associated with high metastatic ability and transfection of CD44 suppresses metastasis without affecting tumorigenecity of rat prostate cancer cells (Gao A C. Lou W. Dong J T. Isaacs J T. CD44 is a metastasis suppressor gene for prostatic cancer located on human chromosome 11p13. Cancer Research. 57(5):846-9. 1997). CD44 down-regulation is a prognostic marker for prostate cancer (Noordzij M A. van Steenbrugge G J. Verkaik N S. Schroder F H. van der Kwast T H. The prognostic value of CD44 isoforms in prostate cancer patients treated by radical prostatectomy. Clinical Cancer Research. 3(5):805-15, 1997).
Human prostate cancer histology is heterogeneous, or xe2x80x9cmulti-focalxe2x80x9d in nature (Isaacs J T. Bova G S. Prostate Cancer. In The Genetic Basis of Human Cancer (Eds. Vogelstein B and Kinzler K W), pp653-60, McGraw-Hill Health Professions Division, 1998 and Bova, 1998), with an average of five lesions in a patient (Bastacky S I. Wojno K J. Walsh P C. Carmichael M J. Epstein J I. Pathological features of hereditary prostate cancer. Journal of Urology. 153(3 Pt 2):987-92, 1995). Between the tumor regions of a prostate and within a single tumor region, the genetic causes to the cancer are heterogeneous and independent as well (Sakr W A. Macoska J A. Benson P. Grignon D J. Wolman S R. Pontes J E. Crissman J D. Allelic loss in locally metastatic, multisampled prostate cancer. Cancer Research. 54(12):3273-7, 1994; Qian J. Bostwick D G. Takahashi S. Borell T J. Herath J F. Lieber M M. Jenkins R B. Chromosomal anomalies in prostatic intraepithelial neoplasia and carcinoma detected by fluorescence in situ hybridization. Cancer Research. 55(22):5408-14, 1995; Mirchandani D. Zheng J. Miller G J. Ghosh A K. Shibata D K. Cote R J. Roy-Burman P. Heterogeneity in intratumor distribution of p53 mutations in human prostate cancer. American Journal of Pathology. 147(1):92-101, 1995). In the end, the best predictor of prostate cancer""s potential to gain invasiveness may be a consideration of a number of genes whose expression levels change along the course of cancer development, as demonstrated in principle by Greene et al. (Greene G F. Kitadai Y. Pettaway C A. von Eschenbach A C. Buucana C D. Fidler I J. Correlation of metastasis-related gene expression with metastatic potential in human prostate carcinoma cells implanted in nude mice using an in situ messenger RNA hybridization technique. American Journal of Pathology. 150(5):1571-82, 1997). Hence, expanding the repertory of such genes, including both the onco-genes and the suppressor genes, will enhance the accuracy and dependability of this approach.
As for the treatment options of prostate cancer, patients with truly organ-confined prostate cancer can be cured by radical prostatectomy. Patients with non-organ-confined prostate cancers, however, have very low survival rates and the current treatments are largely ineffective (Yagoda A. Petrylak D. Cytotoxic chemotherapy for advanced hormone-resistant prostate cancer. [Review] [63 refs] Cancer. 71(3 Suppl):1098-109, 1993).
Breast cancer is the most diagnosed cancer in women and the second leading cancer related cause of death in women (Greenlee R T, Murray T, Bolden S, Wingo P A. Cancer statistics, 1999. Ca: a Cancer Journal for Clinicians 2000;50:7-33). Breast ductal carcinoma in situ (hereinafter indicated as DCIS) incidence has increased dramatically since 1983 as a result of implementing screening programs (Ernster V L. Barclay J. Increases in ductal carcinoma in situ (DCIS) of the breast in relation to mammography: a dilemma. [Review] [34 refs] Journal of the National Cancer Institute. Monographs. (22):151-6, 1997). DCIS is described as a malignant growth of epithelial cells within the ducts and lobules of the breast, and is believed to be the precursor of all invasive breast carcinoma. DCIS itself is non-life-threatening; however, current treatment options for DCIS include masectomy, lumpectomy, radiotherapy or tamoxifen (Ernster V L. Barclay J. Increases in ductal carcinoma in situ (DCIS) of the breast in relation to mammography: a dilemma. [Review] [34 refs] Journal of the National Cancer Institute. Monographs. (22):151-6, 1997; Hwang E S. Esserman L J. Management of ductal carcinoma in situ. [Review] [91 refs] Surgical Clinics of North America. 79(5):1007-30, viii, 1999). These treatment options for DCIS are at best controversial due primarily to a lack of precision in diagnosis and prognosis of whether the detected DCIS will progress to invasive breast cancer and whether recurrence is likely after treatment, usually with a high percentage being invasive breast cancer (Zaugg K. Bodis S. Is there a role for molecular prognostic factors in the clinical management of ductal carcinoma in situ (DCIS) of the breast? [Review] [49 refs] Radiotherapy and Oncology. 55(2):95-9, 2000). At present, histological grading (nuclear grading and whether come do-type necrosis is present) and the size of the DCIS are used to provide assessments of risk of DCIS to progress into invasive breast cancer (Zaugg K. Bodis S. Is there a role for molecular prognostic factors in the clinical management of ductal carcinoma in situ (DCIS) of the breast? [Review] [49 refs] Radiotherapy and Oncology. 55(2):95-9, 2000; Shoker B S. Sloane J P. DCIS grading schemes and clinical implications. [Review] [40 refs] Histopathology. 35(5):393-400, 19; van de Vijver M J. Ductal carcinoma in situ of the breast: histological classification and genetic alterations. [Review] [69 refs] Recent Results in Cancer Research. 152:123-34, 1998). These parameters are far from being adequate for making the most accurate choice of treatment, resulting in a choice either overly excessive or conservative, in either case, the patient will suffer unnecessarily. Molecular markers can help improve our ability to better diagnose DCIS and stratify treatment options, especially the molecular markers that, themselves, play a role in the progression of DCIS to invasive breast cancer (Zaugg K. Bodis S. Is there a role for molecular prognostic factors in the clinical management of ductal carcinoma in situ (DCIS) of the breast? [Review] [49 refs] Radiotherapy and Oncology. 55(2):95-9, 2000; Silverstein M J. Masetti R. Hypothesis and practice: are there several types of treatment for ductal carcinoma in situ of the breast? [Review] [53 refs] Recent Results in Cancer Research. 152:105-22, 1998). The tumorigenesis process is a multi-step transformation in which molecular events escalate to the final stage of invasive phenotype (Silverstein M J. Masetti R. Hypothesis and practice: are there several types of treatment for ductal carcinoma in situ of the breast? [Review] [53 refs] Recent Results in Cancer Research. 152:105-22, 1998). The conventional paradigm of protease involvement in the development and progression of cancer has been the assignment of a usually negative role to the proteases, such as promoting tumor invasion (Mignatti P. Rifkin D B. Biology and biochemistry of proteinases in tumor invasion. [Review] [306 refs] Physiological Reviews. 73(1):161-95, 1993). In turn, the conventional paradigm of protease inhibitors in relation to cancer is usually regard of a beneficial effect for the presence of these molecules (Kennedy A R. Chemopreventive agents: protease inhibitors. Pharmacol Therapeut 78:167-209, 1998). Recently, however, the picture of a new paradigm is beginning to emerge for several serine proteases in breast, prostate, and testicular cancers. A xe2x80x9cnormal epithelial cell specific-1xe2x80x9d (NES1) serine protease was found to be down-regulated in breast and prostate cancers, and it functions as a tumor suppressor (Goyal J, Smith K M, Cowan J M, Wazer D E, Lee S W, Band V. The role for NES1 serine protease as a novel tumor suppressor. Cancer Res 58:4782-4786, 1998). A prostate-specific serine protease, prostase (Nelson P S. Gan L. Ferguson C. Moss P. Gelinas R. Hood L. Wang K. Molecular cloning and characterization of prostase, an androgen-regulated serine protease with prostate-restricted expression. Proceedings of the National Academy of Sciences of the United States of America. 96(6):3114-9, 1999), was shown to be expressed in normal prostate but not in prostate cancer cell lines DU-145 and PC-3. The expression of a testis-specific serine protease, testisin, was shown to be lost in testicular cancer through either a loss of gene (Hooper J D. Nicol D L. Dickinson J L. Eyre H J. Scarman A L. Normyle J F. Stuttgen M A. Douglas M L. Loveland K A. Sutherland G R. Antalis T M. Testisin, a new human serine proteinase expressed by premeiotic testicular germ cells and lost in testicular germ cell tumors. Cancer Research. 59(13):3199-205, 1999) or methylation in the promoter (Boucaut K, Douglas M, Clements J, Antalis T. The serine proteinase testisin may act as a tumor and/or growth suppressor in the testis and may be regulated by DNA methylation. Cancer Genetics and Tumor Suppressor Genes, Cold Spring Harbor Laboratory, 2000). Further, transfection of human testicular cancer cells with a testisin cDNA reduced the tumor growth of xenografts of these cells in nude mice, suggesting a tumor suppressor function for testisin (Boucaut K, Douglas M, Clements J, Antalis T. The serine proteinase testisin may act as a tumor and/or growth suppressor in the testis and may be regulated by DNA methylation. Cancer Genetics and Tumor Suppressor Genes, Cold Spring Harbor Laboratory, 2000). The testisin serine protease is potentially membrane-bound as suggested by its structure and confirmed by immunohistochemistry gene (Hooper J D. Nicol D L. Dickinson J L. Eyre H J. Scarman A L. Normyle J F. Stuttgen M A. Douglas M L. Loveland K A. Sutherland G R. Antalis T M. Testisin, a new human serine proteinase expressed by premeiotic testicular germ cells and lost in testicular germ cell tumors. Cancer Research. 59(13):3199-205, 1999.) Prostasin serine protease is an acidic protein (pI 4.5-4.8) of approximately 40 kDa in molecular mass (Yu J X. Chao L. Chao J. Prostasin is a novel human serine proteinase from seminal fluid. Purification, tissue distribution, and localization in prostate gland. Journal of Biological Chemistry. 269(29):18843-8, 1994). It is predominantly made in the prostate gland (xcx9c140 ng/mg protein), with lesser amounts (2-6 ng/mg protein) also found in the bronchi, colon, kidney, liver, lung, pancreas, and the salivary glands (Yu J X. Chao L. Chao J. Prostasin is a novel human serine proteinase from seminal fluid. Purification, tissue distribution, and localization in prostate gland. Journal of Biological Chemistry. 269(29):18843-8, 1994). Prostasin is secreted in the prostatic fluid, and can be detected in the semen (xcx9c9 xcexcg/ml). Prostasin expression is localized to the epithelial cells of human prostate gland by in situ hybridization histochemistry using an antibody or an anti-sense RNA probe (Yu J X. Chao L. Chao J. Prostasin is a novel human serine proteinase from seminal fluid. Purification, tissue distribution, and localization in prostate gland. Journal of Biological Chemistry. 269(29):18843-8, 1994; (Yu J X. Chao L. Chao J. Molecular cloning, tissue-specific expression, and cellular localization of human prostasin mRNA. Journal of Biological Chemistry. 270(22):13483-9, 1995). Molecular cloning of a full-length human prostasin cDNA revealed that its predicted amino acid residue sequence contains a carboxyl-terminal hydrophobic region that can potentially anchor the protein on the membrane (Yu J X Chao L. Chao J. Molecular cloning, tissue-specific expression, and cellular localization of human prostasin mRNA. Journal of Biological Chemistry. 270(22):13483-9, 1995). At the amino acid level, prostasin is similar to plasma kallikrein, coagulation factor XI, hepsin, plasminogen, acrosin, prostase, and, in particular, testisin (sharing 44% sequence identity) [Nelson P S. Gan L. Ferguson C. Moss P. Gelinas R. Hood L. Wang K. Molecular cloning and characterization of prostase, an androgen-regulated serine protease with prostate-restricted expression. Proceedings of the National Academy of Sciences of the United States of America. 96(6):3114-9, 1999; Hooper J D. Nicol D L. Dickinson J L. Eyre H J. Scarman A L. Normyle J F. Stuttgen M A. Douglas M L. Loveland K A. Sutherland G R. Antalis T M. Testisin, a new human serine proteinase expressed by premeiotic testicular germ cells and lost in testicular germ cell tumors. Cancer Research. 59(13):3199-205, 1999; Yu J X. Chao L. Chao J. Molecular cloning, tissue-specific expression, and cellular localization of human prostasin mRNA. Journal of Biological Chemistry. 270(22): 13483-9, 1995). A membrane-bound Xenopus kidney epithelial cell sodium channel-activating protease (CAP1) was shown to be highly homologous to human prostasin as well (sharing 53% sequence identity at the amino acid level) (Vallet V. Chraibi A. Gaeggeler H P. Horisberger J D. Rossier B C. An epithelial serine protease activates the amiloride-sensitive sodium channel. Nature. 389(6651):607-10, 1997). Prostasin is encoded by a single-copied gene, which is located on human chromosome 16p11.2 (Yu J X. Chao L. Ward D C. Chao J. Structure and chromosomal localization of the human prostasin (PRSS8) gene. Genomics. 32(3):334-40, 1996). The secreted prostasin cleaves synthetic substrates in vitro preferentially at the carboxyl-terminal side of Arg residue, and is thus considered a trypsin-like serine protease (Yu J X. Chao L. Chao J. Prostasin is a novel human serine proteinase from seminal fluid. Purification, tissue distribution, and localization in prostate gland. Journal of Biological Chemistry. 269(29):18843-8, 1994). The physiological function of prostasin, however, has remained unknown. By comparing gene expression of normal tissues, pre-invasive cancer, and invasive cancer, it would be highly advantageous to discover molecular markers that display a differential expression pattern between the pre-invasive and the invasive phenotypes and use these markers for a more precise diagnosis and prognosis of prostate and/or breast cancers. With the enhanced precision in diagnosis and prognosis, treatment options for patients with DCIS could then be stratified. Those with low risks of developing invasive breast cancer will have a higher confidence in choosing breast-conserving options, while those at high risks will ponder more aggressive options with the necessary follow-up treatments. Similarly, those males with low risks of developing invasive prostate cancer will have a higher confidence in choosing prostate-conserving options, while those at high risks will ponder more aggressive options with the necessary follow-up treatments.
The first objective of the present invention is to reduce deficiencies in the prior art with specific regard to differential diagnosis of invasive prostate and breast cancers and to treatment of invasive and metastatic prostate and breast cancers.
The second objective of the present invention is to provide a new marker for prostate and breast cancer.
The third objective of the invention is to provide as a drug to patients with carcinoma of the prostate via delivery of a functional prostasin gene.
The fourth objective of the invention is to provide as a drug to patents with carcinoma of the prostate via delivery of a functional prostasin cDNA.
The fifth objective of the invention is to provide as a drug to patents with carcinoma of the breast via delivery of a functional prostasin gene.
The sixth objective of the invention is to provide as a drug to patents with carcinoma of the breast via delivery of a functional prostasin cDNA.
This invention identifies prostasin serine protease as a potential marker, and as a tumor invasion suppressor for prostate and breast cancers and thus provides methods (a), (b), and (c) of determining invasiveness levels of human carcinomas:
(a) using prostasin protein levels, comprising the steps of: sampling a human carcinoma tissue; determining prostasin protein levels in the human carcinoma tissue; preferably by applying an immunological reagent-antibody to said tissue wherein the reagent-antibody becomes bound to prostasin protein in said tissue, and determining invasiveness of the human carcinoma tissue based on the prostasin protein levels; or
(b) using prostasin mRNA levels, comprising the steps of: sampling a human carcinoma tissue; determining prostasin mRNA levels in the human carcinoma tissue preferably by applying prostasin-specific anti-sense RNA probes in an in situ hybridization to determine the prostasin mRNA levels in the separated human carcinoma tissue the determination of the prostasin mRNA levels in the separated human carcinoma sample tissue; to make possible and determining invasiveness of the human carcinoma tissue based on the prostasin mRNA levels; or
(c) (c) using prostasin gene promoter DNA methylation levels, comprising the steps of: sampling a human carcinoma tissue; determining prostasin gene promoter DNA methylation levels in the human carcinoma tissue, preferably by applying prostasin-promoter-specific oligonucleotide primers in a PCR to determine the prostasin gene promoter DNA methylation levels in the sampled human carcinoma tissue and determining invasiveness of the human carcinoma tissue based on the prostasin gene promoter DNA methylation levels,
as well as a method of treating invasive human carcinomas comprising the steps of: incorporating human prostasin nucleic acid into a selected gene delivery vector nucleic acid to form a recombinant nucleic acid; preferably wherein the nucleic acid includes a gene or cDNA delivering the recombinant nucleic acid into a human carcinoma; and reducing invasiveness of the human carcinoma.